Rotational Diffusion of Intrinsically Disordered Proteins Under Crowded Conditions
Juan Armas, Layla Chirar, Lika Chhit, Maureen Cabrera Montes, Evan Forrest
Department of Chemistry & Biochemistry
Faculty Supervisor: Raymond Esquerra
Tau and α-synuclein (α-syn) are intrinsically disordered proteins (IDPs) implicated in Alzheimer’s and Parkinson’s diseases through misfolding and aggregation. Their lack of stable structure makes them highly sensitive to environmental conditions, including temperature, molecular interactions, and macromolecular crowding. However, most in vitro studies are performed in dilute buffers that do not reflect the crowded, viscous intracellular environment where aggregation occurs.
To better approximate cellular conditions, we introduce chemically inert macromolecular crowding agents and examine their effects on IDP conformational dynamics and aggregation. We employ a myoglobin fusion strategy to enable time-resolved linear dichroism (TRLD) measurements, allowing extraction of rotational correlation times as a readout of microviscosity and conformational restriction. Complementary techniques, including thioflavin fluorescence and circular dichroism spectroscopy, are used to assess structural changes and aggregation behavior. By linking intracellular-like physical constraints to rotational diffusion and aggregation propensity, this work aims to provide mechanistic insight into how cellular environments influence disease-relevant IDP behavior.